Ex vivo modelling of PD-1/PD-L1 immune checkpoint blockade under acute, chronic, and exhaustion-like conditions of T-cell stimulation.

Blockade of PD-1/PD-L1 interactions is proving an exciting, durable therapeutic modality in a range of cancers whereby T cells are released from checkpoint inhibition to revive their inherent anti-tumour activity. Here we have studied various ways to model ex vivo T cell function in order to compare the impact of the clinically utilised anti-PD-1 antibody, pembrolizumab (Keytruda) on the activation of human T cells: focussing on the release of pro-inflammatory IFNγ and anti-inflammatory IL-10 to assess functionality. Firstly, we investigated the actions of pembrolizumab in an acute model of T-cell activation with either immature or mature allogeneic dendritic cells (DCs); pembrolizumab enhanced IFNγ and IL-10 release from purified CD4+ T-cells in the majority of donors with a bias towards pro-inflammatory cytokine release. Next, we modelled the impact of pembrolizumab in settings of more chronic T-cell activation. In a 7-day antigen-specific response to EBV peptides, the presence of pembrolizumab resulted in a relatively modest increase in both IFNγ and IL-10 release. Where pembrolizumab was assessed against long-term stimulated CD4+ cells that had up-regulated the exhaustion markers TIM-3 and PD-1, there was a highly effective enhancement of the otherwise exhausted response to allogeneic DCs with respect to IFNγ production. By contrast, the restoration of IL-10 production was considerably more limited. Finally, to assess a direct clinical relevance we investigated the consequence of PD-1/PD-L1 blockade in the disease setting of dissociated cells from lung and colon carcinomas responding to allogeneic DCs: here, pembrolizumab once more enhanced IFNγ production from the majority of tumour preparations whereas, again, the increase in IL-10 release was modest at best. In conclusion, we have shown that the contribution of PD-1-revealed by using a canonical blocking antibody to interrupt its interaction with PD-L1-to the production of an exemplar pro- and anti-inflammatory cytokine, respectively, depends in magnitude and ratio on the particular stimulation setting and activation status of the target T cell. We have identified a number of in vitro assays with response profiles that mimic features of dissociated cell populations from primary tumours thereby indicating these represent disease-relevant functional assays for the screening of immune checkpoint inhibitors in current and future development. Such in vitro assays may also support patient stratification of those likely to respond to immuno-oncology therapies in the wider population.

The evolution of host resistance and parasite infectivity is highest in seasonal resource environments that oscillate at intermediate amplitudes.

Seasonal environments vary in their amplitude of oscillation but the effects of this temporal heterogeneity for host-parasite coevolution are poorly understood. Here, we combined mathematical modelling and experimental evolution of a coevolving bacteria-phage interaction to show that the intensity of host-parasite coevolution peaked in environments that oscillate in their resource supply with intermediate amplitude. Our experimentally parameterized mathematical model explains that this pattern is primarily driven by the ecological effects of resource oscillations on host growth rates. Our findings suggest that in host-parasite systems where the host's but not the parasite's population growth dynamics are subject to seasonal forcing, the intensity of coevolution will peak at intermediate amplitudes but be constrained at extreme amplitudes of environmental oscillation.

The effect of temporal fluctuations on the evolution of host tolerance to parasitism.

There are many mechanisms that hosts can evolve to defend against parasites, two of which are resistance and tolerance. These defences often have different evolutionary behaviours, and it is important to consider how each individual mechanism may respond to changes in environment. In particular, host defence through tolerance is predicted to be unlikely to lead to variation, despite many observations of diversity in both animal and plant systems. Hence understanding the drivers of diversity in host defence and parasite virulence is vital for predicting future evolutionary changes in infectious disease dynamics. It has been suggested that heterogeneous environments might generally promote diversity, but the effect of temporal fluctuations has received little attention theoretically or empirically, and there has been no examination of how temporal fluctuations affects the evolution of host tolerance. In this study, we use a mathematical model to investigate the evolution of host tolerance in a temporally fluctuating environment. We show that investment in tolerance increases in more variable environments, giving qualitatively different evolutionary behaviours when compared to resistance. Once seasonality is introduced evolutionary branching though tolerance can occur and create diversity within the population, although potentially only temporarily. This branching behaviour arises due to the emergence of a negative feedback with the maximum infected density on a cycle, which is strongest when the infected population is large. This work reinforces the qualitative differences between tolerance and resistance evolution, but also provides theoretical evidence for the theory that heterogeneous environments promote host-parasite diversity, hence constant environment assumptions may omit important evolutionary outcomes.

The evolution of host defence to parasitism in fluctuating environments.

Given rapidly changing environments, it is important for us to understand how the evolution of host defence responds to fluctuating environments. Here we present the first theoretical study of evolution of host resistance to parasitism in a classic epidemiological model where the host birth rate varies seasonally. We show that this form of seasonality has clear qualitative and quantitative impacts on the evolution of resistance. When the host can recover from infection, it evolves a lower level of defence when the amplitude is high. However, when recovery is absent, the host increases its defence for higher amplitudes. Between these different behaviours we find a region of parameter space that allows evolutionary bistability. When this occurs, the level of defence the host evolves depends on initial conditions, and in some cases a switch between attractors can lead to different periods in the population dynamics at each of the evolutionary stable strategies. Crucially, we find that evolutionary behaviour found in a constant environment for this model doesn't always hold for hosts with highly variable birth rates. Hence we argue that seasonality must be taken into account if we want to make predictions about evolutionary trends in real-world host-parasite systems.